HIPK2 is a member of the recently identified family of nuclear protein kinases (HIPK1, HIPK2 and HIPK3) that localizes to nuclear dots by SUMO-1 modification (sumoylation) and is a component of co-repressor complex recruited by the NK-3 homeodomain protein. Using confocal microscopy, co-immunoprecipitation, in vitro pull-down assays and chemical cross linking experiments we show that HIPK2 can form a dimer with either HIPK1 or HIPK2. The dimerization domain maps to the carboxyl-terminal half of HIPK2. Forced expression of either the kinase-inactive form of the full-length HIPK2 or a truncated form of the carboxyl-terminal half of HIPK2 in zebrafish embryos disrupted early embryogenesis whereas the expression of the amino-terminal half of HIPK2 including the active kinase domain caused the one-eyed phenotype. Thus, theses results suggest that the dimer formation among HIPKs may have a pivotal role in regulation of HIPK2 function during development. We have previously demonstrated that HIPK2 exclusively localizes to nuclear dots by sumoylation. Research in progress shows that the sumoylated HIPK2 is a target for the SUMO-specific protease SENP-1. Cotransfection of SENP-1 remarkably reduced sumolyated HIPK2 and formation of the HIPK2-positive nuclear dots, implying that desumoylation of HIPK2 regulates dissociation of HIPK2 from nuclear dots. Interestingly, SENP-1 shuttles between nucleus and cytoplasm, revealed by treatment of transfected cells with Leptomycin B, an inhibitor of nuclear export of proteins. Search for signal transduction pathways that regulate SUMO-modification of HIPK2 is underway. cDNAs for the Drosophila HIPK2 have been cloned and sequenced. In contrast to the HIPK family in mammals there is only one hipk gene (we named it Dhipk2) in the Drosophila genome. However, northern blot analysis reveals multiple Dhipk2 transcripts. P-element insertion lines that cause embryonic lethality has also been isolated. Genetic analysis of embryos from these flies suggests that Dhipk2 is involved in multiple developmental pathways. Of note, expression of HIPK2 in cardial cells causes abnormal positioning of the dorsal vessel, suggesting that DHIPK2 may play a role in cardiac development.